This project is exploring structure-function relationships involved in activation and assembly of the microbicidal NADPH oxidase of phagocytes (the respiratory burst). Earlier work has defined four essential oxidase components (p47-phox, p67-phox, p22-phox and gp91-phox) as proteins absent or defective in various forms of chronic granulomatous disease (CGD), a genetic disease characterized by enhanced susceptibility to bacterial and fungal pathogens. Our current efforts are studying mechanisms of regulation of NADPH oxidase and exploring methods for inhibiting NADPH oxidase function by disrupting interactions among oxidase components. We have recently presented a model for oxidase assembly involving multiple SH3 domain interactions between oxidase components. We have now identified two factors that down-regulate oxidase activity by affecting SH3 domain interactions. p40phox disrupts the oxidase by competing with the tail-tail SH3 interaction between p67phox and a proline-rich target in p47phox. Pak is a Rac-regulated protein kinase that binds to the first SH3 domain of p47phox and down-regulates the oxidase in a phosphorylation-dependent manner. In other work we have identified several proline-rich, SH3-binding ligands from combinatorial peptide phage display libraries. These peptides all bind to the first SH3 domain of p47phox and could serve as specific inhibitors of the oxidase. Other work in progress is exploring the roles of several other protein kinases and phospholipases as potential targets for modulating oxidase activity. These studies will provide insights on this important oxygen-dependent host defense system and may aid in the design of drugs that inhibit production of reactive oxygen species, which are thought to have a role in inflammatory disease processes.